The long-term interests of this project are to determine the neurohormonal mechanisms that contribute to the ability of the HPA axis to habituate to repeated stress. Understanding the mechanisms of the habituation of the HPA axis stress response may lead to a better understanding of the mechanisms underlying dysregulation of the HPA axis that is present in a large number of individuals with clinical depression. A strong case has been made for viewing depression as a stress-related disorder, and at a physiological level depression may be a maladaptive response to repeated stress in which sensitization predominates, perhaps because of an impairment in habituation mechanisms. The operation of the HPA axis is extremely sensitive to corticosteroid feedback inhibition, and one means by which repeated stress may modify the HPA axis is by producing changes at the corticosteroid receptor level. A widely held hypothesis is that the increased level of corticosteroids present during chronic stress produces corticosteroid receptor downregulation, resulting in additional HPA axis hyperactivity. While some repeated stress studies have provided evidence for corticosteroid receptor downregulation in the hippocampus of the rat brain, other studies have failed to observe decreases in corticosteroid receptors with repeated stress, and on some occasions, increases in corticosteroid receptors were observed. The discrepancies in stress- induced corticosteroid receptor changes may pertain to whether the stress regimen is one that results in sensitization or habituation of the HPA axis stress response. For example, severe chronic stress may produce a decrease in corticosteroid receptor level and lead to increased HPA axis activity (sensitization), whereas a milder repetitive stressor may produce an increase in corticosteroid receptors and lead to decreased HPA axis activity (habituation). This latter prospect has been largely unexplored, and will be the focus of this project. Past studies examining in vivo stress effects on corticosteroid receptor levels in the brain have been hindered by the limitations of the cytosolic receptor binding assay, that until now has been the best means available for quantitatively measuring corticosteroid receptors. This homogenate based receptor binding procedure may not provide the resolution that is necessary to detect discrete changes in corticosteroid receptor levels within subregions of brain tissue. It is anticipated that studies described in this proposal will be able to overcome this limitation by the use of a radioimmunohistochemical method for measuring relative corticosteroid receptor concentrations in rat brain tissue sections. The following two specific aims are proposed: Specific Aim 1. To further validate the use of a radioimmunohistochemical procedure to measure relative changes in corticosteroid receptor levels in pituitary and brain tissue. Specific Aim 2. To use radioimmunohistochemistry to determine if there are regional changes in Type I or Type II corticosteroid receptor levels as a consequence of repeated restraint stress.